Engineering mediator-based electroactivity in the obligate aerobic bacterium Pseudomonas putida KT2440.
Engineering mediator-based electroactivity in the obligate aerobic bacterium Pseudomonas putida KT2440.
复制标题
DOI：
10.3389/fmicb.2015.00284
复制
发表时间：
2015
影响因子：
5.2
通讯作者：
Rosenbaum MA
中科院分区：
文献类型：
--
作者：
Schmitz S;Nies S;Wierckx N;Blank LM;Rosenbaum MA
恶臭假单胞菌菌株正在被开发为微生物生产宿主，用于生产一系列两亲性和疏水性生化产品。因此，恶臭假单胞菌的专性需氧生长可能是经济和技术上的挑战，因为它需要持续严格的通气并且经常导致反应器起泡。在这里，我们设计了一种恶臭假单胞菌 KT2440 菌株，它可以从铜绿假单胞菌中产生吩嗪氧化还原介体，从而在限氧条件下与电极实现部分氧化还原平衡。已知铜绿假单胞菌利用其吩嗪型氧化还原介体与生物电化学系统（BES）中的阳极进行电子交换。我们将两个诱导型质粒上的七个核心吩嗪生物合成基因 phzA-G 以及铜绿假单胞菌绿脓素合成所需的两个特定基因 phzM 和 phzS 转移到恶臭假单胞菌 KT2440 中。最好的克隆 P. putida pPhz 在 25 小时的生长过程中产生了 45 mg/L 的绿脓素，形成蓝色可见，与铜绿假单胞菌的绿脓素产量相当。然后，通过电化学氧化还原控制，在不同的限氧条件下对这种新菌株进行了表征，并与未修饰的恶臭假单胞菌 KT2440 相比，评估了中心能量代谢的变化。在新菌株中，上清液浓度高达 33 μg/mL 的吩嗪合成与向阳极释放电子的能力呈线性相关，其中吩嗪-1-羧酸充当主要的氧化还原介体。 P. putida pPhz 在高达 12 μA/cm2 的阳极电流密度下可维持强烈的限氧代谢长达 2 周。总之，这项工作为未来恶臭假单胞菌菌株的限氧生物催化奠定了基础。
Pseudomonas putida strains are being developed as microbial production hosts for production of a range of amphiphilic and hydrophobic biochemicals. P. putida's obligate aerobic growth thereby can be an economical and technical challenge because it requires constant rigorous aeration and often causes reactor foaming. Here, we engineered a strain of P. putida KT2440 that can produce phenazine redox-mediators from Pseudomonas aeruginosa to allow partial redox balancing with an electrode under oxygen-limited conditions. P. aeruginosa is known to employ its phenazine-type redox mediators for electron exchange with an anode in bioelectrochemical systems (BES). We transferred the seven core phenazine biosynthesis genes phzA-G and the two specific genes phzM and phzS required for pyocyanin synthesis from P. aeruginosa on two inducible plasmids into P. putida KT2440. The best clone, P. putida pPhz, produced 45 mg/L pyocyanin over 25 h of growth, which was visible as blue color formation and is comparable to the pyocyanin production of P. aeruginosa. This new strain was then characterized under different oxygen-limited conditions with electrochemical redox control and changes in central energy metabolism were evaluated in comparison to the unmodified P. putida KT2440. In the new strain, phenazine synthesis with supernatant concentrations up to 33 μg/mL correlated linearly with the ability to discharge electrons to an anode, whereby phenazine-1-carboxylic acid served as the dominating redox mediator. P. putida pPhz sustained strongly oxygen-limited metabolism for up to 2 weeks at up to 12 μA/cm2 anodic current density. Together, this work lays a foundation for future oxygen-limited biocatalysis with P. putida strains.
登录
查看更多内容
影响因子：
6.4
作者：
Flynn, Jeffrey M.;Ross, Daniel E.;Gralnick, Jeffrey A.
通讯作者：
Gralnick, Jeffrey A.
影响因子：
1.9
作者：
Kuepper, Benjamin;Mause, Anika;Wichmann, Rolf
通讯作者：
Wichmann, Rolf
影响因子：
48
作者：
Gibson, Daniel G.;Young, Lei;Smith, Hamilton O.
通讯作者：
Smith, Hamilton O.
影响因子：
5.4
作者：
Blank, Lars M.;Ionidis, Georgios;Schmid, Andreas
通讯作者：
Schmid, Andreas
影响因子：
3.2
作者：
Mavrodi, DV;Bonsall, RF;Thomashow, LS
通讯作者：
Thomashow, LS